Duchenne muscular dystrophy (DMD) is a common X-linked recessive disease within the male population. DMD is characterized by a slow, irreversible deterioration of skeletal muscle, and few patients survive past their second decade. To date, neither the genetic defect nor the biochemical mechanism of this disease has been determined. The goal of this proposal is to follow two separate leads that promise to elucidate the molecular basis of DMD. Human blood platelets (of normal and DMD individuals) will be studied. For our first lead, we have tentatively observed that DMD platelets respond abnormally to certain aggregating agents (e.g. epinephrine and collagen). We propose to extend our aggregation analyses by using both a larger patient population and an expanded repertoire of aggregating agents. The protocol also includes the quantitation of platelet granule secretion. The initiation of platelet secretion and aggregation involves the activation of several discrete protein kinases. Therefore, the experimental protocol will focus on the phosphorylation of polypeptides during the induction of secretion and aggregation by agents that can selectively activate various protein kinase pathways. These studies, using 2-D gel and autoradiographic analyses, seek to define differences in important enzymatic processes between normal and DMD platelets. For our second lead, we have observed high molecular differences in the 2-D gel polypeptide patterns of whole platelet extracts when comparing normal and DMD platelets. The cytoskeletal association of two high molecular weight polypeptides, myosin and actin-binding protein (ABP), seems to be involved in the normal secretion and/or aggregation. Therefore, we propose to analyze 2-D gel patterns of platelet cytoskeleton, myosin, and ABP before and after the exposure of platelets to various aggregating agents. Myosin heavy chain, ABP, and the 'abnormal' high molecular weight polypeptide(s) will be analyzed by peptide mapping. These studies seek to find potential disease-related difference in cytoskeletal polypeptide associations as well as differences in high molecular weight polypeptides associations as well as differences in high molecular weight polypeptides. Combined with the protein kinase studies, gel patterns of contractile polypeptide preparations may characterize the critical difference between normal and DMD platelets.